Blood and saliva samples of 122 like-sexed twin pairs (65 MZ and 57 DZ) living in Chandigarh (India) were analyzed for ABH polymorphism. The results indicated that ABH secretions were independent of ABO blood groups though there was an indication of higher incidence of non-secretors among ‘O’ blood group twin individuals. No significant differences were observed between twins and singletons in secretor gene frequency estimates. The quantitative data revealed that mean titre scores for H substances were lower than that for A and B substances. F test contrasting intra-pair variance between zygosities for ABH quantitative secretions was highly significant indicating stronger genetic component of variation. The results suggested that quantitative assay of ABH secretions would be a better indicator for zygosity determination than mere qualitative differentiation.

Parents of a sample of 76 same sexed pairs of twins aged 3 to 9 months completed a mailed similarity questionnaire. It included the Bonnelykke et al.'s questionnaire and a four anthropological variable scale. To improve each of these two methods, three other combined methods were carried out and results were compared with the biological zygosity diagnosis. The Bonnelykke et al.'s classification combined with anthropological scale (method 4) gave only 1.2% misclassified in the whole sample. It is concluded that zygosity diagnosis using this type of procedure to distinguish MZ and DZ pairs would be important not only for epidemiological study but also for pediatricians and parents.

Subjects were 74 twin pairs, 61 MZs and 13 same-sexed DZs who entered the High School affiliated with Tokyo University, Japan. Their mothers also participated. The twins’ zygosity was previously identified by many genetic markers. This study aimed at clarifying the effectiveness of zygosity diagnosis by questionnaires distributed to twins’ mothers. The questionnaire consisted of three questions concerning the degree of similarity of twins at one year of age; whether they were confused, and if so, by whom. It was slightly modified from that reported earlier by Ooki et al [11] for twins themselves. According to the degree of similarity of the twins, points were allotted thus: from 1 to 3 points for answers to questions (1) and (2) and from 1 to 4 points for answers to questions (3). The sum of the points was calculated, then ranged from 3 to 10. Zygosity was determined by the sum of these points. If the sum was 3-6, the twin pairs were considered MZ and if the sum was 7-10 DZ. It was revealed that more than 90% of twins were identified correctly as MZ or DZ by applying this cutting point and this result was in accordance with that obtained by use of discriminant function analysis. Moreover, the accuracy of the mothers’ responses was nearly the same as that obtained by the questionnaire for twins themselves. It was concluded, therefore, that the information from twins’ mothers is as correct as that from twins themselves. This questionnaire is simple, practical and especially useful when twins are still too young to participate personally.

Although numerous genetic and anthropological markers are available for determining zygosity of twins, there is still a need for a more practical and informative method in zygosity diagnosis. Dinucleotide repeats or other short repeats (microsatellites) are highly variable between individuals and offer a simple, fast, cheap, and exact approach for zygosity determination. The feasibility of a set of microsatellites to be used for this purpose is demonstrated.

The usual assumption that monochorionic twins are monozygotic has recently been questioned, based on blood group discordance in 3 of 12 monochorionic pairs. Therefore, this study evaluates the validity of zygosity diagnosis based on examination of placental membranes, and at the same time evaluates Weinberg's differential rule in a Danish consecutive twin series. All twin pairs, 265 like-sexed and 87 unlike-sexed pairs, born at Odense University Hospital, Denmark, from 1 January 1980 through 31 August 1988, were ascertained. In 229 like-sexed pairs, zygosity was determined by serological methods, and in 190 of these, the fetal membranes could be reliably classified. Fiftynine monochorionic twin pairs were identified and none of those pairs were discordant for any genetic markers. Therefore, we conclude that monochorionic twins can be considered monozygotic. The distribution of monozygotic and dizygotic pairs was in accordance with Weinberg's rule.

Subjects were 189 twin pairs, 165 MZ and 24 same-sexed DZ, who entered the junior high school affiliated to Tokyo University (sample T), and 93 twin pairs, 71 MZ and 22 same-sexed DZ, who were registered at Kinki University (sample K). The zygosity was previously identified by many genetic markers, and this study aimed at zygosity diagnosis by questionnaire. The latter included three questions: “How are you alike?”, “How often are you mistaken?”, and “By whom are you mistaken?”. According to the degree, 1-3 points, 1-3 points, and 1-4 points were given for each question, and the sum of the points of each pair of twins was calculated. Zygosity was determined by the sum of points, distributed from 6 to 20. Namely, if the sum was 6-13, the twin pair was considered MZ, and if the sum was 14-20, DZ. More than 90% of twins were diagnosed correctly by use of this cutting point. This result was in accordance with that obtained by use of discriminant function analysis. It is concluded that zygosity diagnosis by questionnaire is convenient and useful, in particular for epidemiological research.